PROJECT SUMMARY/ABSTRACT All drugs of abuse produce unique interoceptive/subjective (i.e., discriminative stimulus) effects that can impact drug-taking, seeking, and relapse in both clinical and preclinical studies. However, the neural circuitry modulating the interoceptive effects of alcohol has yet to be established. The nucleus accumbens core (AcbC), a region known to mediate alcohol-related behaviors, also plays a central role in modulating the discriminative stimulus effects of alcohol. Therefore, to begin to establish the circuitry modulating the interoceptive effects of alcohol, two brain regions with projections to the AcbC will be the focus of the proposed studies: the insular cortex (IC) and the rhomboid/reuniens ventral midline thalamic nuclei (RhRe). Despite the implicated role of the IC in processing interoceptive cues and responding to alcohol-related cues, its functional role in modulating alcohol-induced interoceptive effects has not been investigated to date. Additionally, the RhRe are proposed to modulate inhibition, behavior flexibility, and motivation, but their role in modulating any drug-related behaviors has yet to be determined. However, our preliminary data demonstrate that chemogenetic inactivation of the IC increases sensitivity to alcohol and pharmacological inactivation of IC and RhRe (independently) results in partial ?alcohol-like? interoceptive effects, thus implicating a functional role for the IC and RhRe in modulating the discriminative stimulus effects of alcohol. Therefore, by investigating the two novel brain regions (IC and RhRe) and their projections to the AcbC, this application proposes to investigate the neural circuitry underlying alcohol-induced interoceptive states. Using drug discrimination methods, rats will be trained to discriminate alcohol vs. water, and we will chemogenetically inactivate the brain regions and their specific incoming AcbC projections to examine whether sensitivity to alcohol is altered. Thus, experiments in Aim 1 will determine whether IC and its projections to the AcbC modulate the discriminative stimulus effects of alcohol. Aim 2 will use a similar approach to determine the role of RhRe and its projections to the AcbC. Together these experiments will demonstrate a functional role for two novel brain regions in modulating the interoceptive effects of alcohol, while identifying potential AcbC-related neural circuitry. By conducting these experiments in parallel, this innovative approach has the ability to inform the alcohol field of regions not previously implicated or understudied while demonstrating the importance and neurobiology underlying alcohol interoceptive states/internal cues, thus helping bridge the clinical and preclinical fields.